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Background: Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI–TOF MS) is proposed for mosquito species identification. The absence of public repositories sharing mass spectra and open-source data analysis pipelines for fingerprint matching to mosquito species limits the widespread use of this technology. The objective of this study was to develop a free open-source data analysis pipeline for Anopheles species identification with MALDI–TOF MS. Methods: Anopheles mosquitoes were captured in 33 villages in Karen (Kayin) state in Myanmar. A subset of 403 specimens was selected for inclusion in either the reference or the test panel (270 and 133 specimens, respectively). Three hundred fifty-nine specimens could be identified with DNA barcodes and were assigned to 21 sensu stricto species and five sibling species pairs or complexes. A total of 3584 mass spectra of the head of these specimens identified with DNA barcoding were acquired and the similarity between mass spectra was quantified using a cross-correlation approach adapted from the published literature. A reference mass spectra database was created using all spectra of the PCR-identified specimens assigned to the reference panel. A simulation experiment was carried out by querying the reference database with the spectra of the test panel to evaluate the performance of species identification with MALDI–TOF MS at varying thresholds of the cross-correlation index for the algorithm to output an identification result and with varying numbers of technical replicates for the tested specimens, considering PCR identification results as the reference. Results: With one spot and a threshold value of −14 for the cross-correlation index on the log scale, the sensitivity was 0.99 [95% credible interval (CrI): 0.98–1.00], the predictive positive value was 0.99 (95% CrI: 0.98–0.99), and the accuracy was 0.98 (95% CrI: 0.97–0.99). It was not possible to directly estimate the sensitivity and negative predictive value because there was no true negative (i.e., queries of species not referenced in the database) in the assessment. Conclusions: The cross-correlation approach can be used to match mass spectral fingerprints to predefined taxa. MALDI–TOF MS is a valuable tool for rapid, accurate, and affordable identification of Anopheles species. [ABSTRACT FROM AUTHOR]

Limnebius exhibits uniform external appearance but variable male genitalia: from differently curved rod shapes in the subgenus Bilimneus to up to seven longitudinally separated folds or appendages in Limnebius s.s.. Among nonsexual traits, two subgenera diverge only in the wing. The aedeagal complexity in Limnebius s.s. is associated with the secondary sexual structures on the abdomen and metatibia; larger body size, which forms two clusters; and male-biased sexual dimorphism. To cluster the subdivisions with phylogenetically unresolved relationships, we examined the coincidence of the traits in both sexes, based on the premise of the independent and monophyletic origin of histologically unsimilar elements. The folds and additional appendages on the median lobe, abdominal paired rows of setae, and protuberances show that the L. parvulus and L. nitidus groups have maximum aedeagal complexity in the L. parvulus and L. nitiduloides subgroups, respectively. The L. parvulus group exhibits a gradual increase in the number of elements of the aedeagi. The histology of connections of the parameres and additional appendages to the median lobe, along with that of the flagellum opening, reveal the subgroups' relationship in the L. nitidus group, as well as a decrease in the complexity of the male genitalia in their recent branches. [ABSTRACT FROM AUTHOR]

The Atlantic blue crab, Callinectes sapidus, is acknowledged as one of the worst invasive alien species in the Mediterranean, impacting biodiversity and human activities. Native to the western Atlantic, it has expanded to European coastal waters since the early 1900s. Despite its ecological and commercial importance, genetic research on this species is limited. Here we show a comprehensive investigation of the genetic variation and evolutionary history in Callinectes sapidus using 667 mitochondrial COI gene sequences. Our dataset encompasses 36 newly generated sequences from previously understudied Mediterranean sites and 631 from worldwide locations obtained from the GenBank public database. Our findings reveal two distinct, but closely related, genetic groups within the species' distribution range, suggesting the occurrence of a potential species complex. Furthermore, in the Mediterranean, low levels of genetic variability were observed except for substantial haplotypic differentiation in Turkish samples. This study depicts the global genetic diversity and evolutionary patterns of Callinectes sapidus, offering new insights into the taxonomic status of the species. [ABSTRACT FROM AUTHOR]

Adaptation to different ecological niches is considered one of the main drivers of species diversification, also in cryptic species complexes, notwithstanding their morphological stasis. We here review all the published information on ecological differentiation within cryptic species complexes within the phylum Rotifera. We found 177 instances of cryptic species identified genetically, published in 101 papers. Only a subset of the papers contained available information on ecological differences and only for two of the 54 known complexes, namely Brachionus calyciflorus s.l. and Brachionus plicatilis s.l., sufficient data were available for analyses of ecological differences. For the B. calyciflorus complex, B. fernandoi consistently occurred at a significantly lower temperature than the other species in the complex; no differences were found for other environmental variables. For the B. plicatilis complex, B. paranguensis occurred in waters with higher pH than the other species; no differences were found for other environmental variables. These results, even if preliminary and based on scattered information, reveal that adaptation to different ecological niches exists also between closely related species. Our aim is to showcase this interesting field of research spurring further detailed studies to focus on the mechanisms of ecological speciation using rotifer cryptic species as a model system. [ABSTRACT FROM AUTHOR]

5S ribosomal DNAs (rDNAs) are arranged in tandem and are often under-represented in genome assemblies. In the present study, we performed a global and in-depth analysis of the 5S rDNAs in the model insect Tribolium castaneum and its closely related species Tribolium freemani. To accomplish this goal, we used our recently published genome assemblies based on Nanopore and PacBio long-read sequencing. Although these closely related species share the 5S rRNA gene sequence with high homology, they show a different organization of the 5S rDNA locus. Analysis of 5S rDNA arrays in T. castaneum revealed a typical tandemly repeated organization characterized by repeat units consisting of the 121 bp long 5S rRNA gene and the 71 bp long nontranscribed spacer (NTS). In contrast, T. freemani showed a much more complex organization of 5S rDNA arrays characterized by two patterns. The first is based on the association of 5S rRNA gene with arrays of a satellite DNA, representing the NTS sequence of the 5S rDNA genes in T. freemani. The second, more complex type is characterized by a somewhat less frequent occurrence of the 5S rRNA gene and its association with longer satellite DNA arrays that are regularly interrupted by Jockey-like retrotransposons. This organization, in which the ribosomal gene is associated with two completely different repetitive elements such as satellite DNAs and retrotransposons, suggests that the 5S rRNA gene, regardless of its crucial function in the genome, could be a subject of extremely dynamic genomic rearrangements. [ABSTRACT FROM AUTHOR]

The description of diagnostic morphological characters and DNA barcoding of fish larvae from nine species of the carangid family are provided from specimens collected during a weekly zooplankton time-series (2016–2017) at Cabo Pulmo National Park, Gulf of California, Mexico. Five nominal species (Caranx sexfasciatus, C. caballus, Naucrates ductor, Selar crumenophthalmus, and Selene peruviana) and three morphotypes of Decapterus spp. and one of Caranx spp. were identified and separated based on morphological, meristic, and pigmentary diagnostic characters. All larvae were genetically sequenced for a fragment of the cytochrome c oxidase subunit I mitochondrial gene. Sequences of larval Caranx and Decapterus showed high genetic similarity (> 99%), low intraspecific divergence (< 1%), and an interspecific divergence between 6% and 11%, allowing the discrimination of diagnostic pigmentation patterns of fish larvae among three sibling species from each genus: Caranx (C. caballus, C. caninus, and C. sexfasciatus) and Decapterus (D. macarellus, D. macrosoma, and D. muroadsi). DNA barcoding supported the presence of Caranx caballus, C. caninus, C. sexfasciatus, Decapterus macarellus, D. muroadsi, Selar crumenophthalmus, and Selene peruviana, and for the first time Naucrates ductor and D. macrosoma at the CPNP. Abundance of these nine species (confirmed molecularly) was estimated throughout the 2016–2017 weekly time series. Decapterus macarellus and Caranx caninus were the most abundant species. The morphological and molecular taxonomic methods allowed us to infer the species number and abundance of these commercial species at the CPNP to improve conservation in protected areas and fishery management. [ABSTRACT FROM AUTHOR]

The widespread Western Australian volute that is referred to as "Cymbiola nivosa" is shown here to actually represent a complex of closely-related sibling species, each with its own distinct geographical distribution: a northern species, Cymbiola oblita (E.A. Smith, 1909), which ranges from near Port Hedland north to Cape Leveque and the Dampier Archipelago; a central species Cymbiola nivosa (Lamarck, 1804), which ranges from Port Hedland south to Shark Bay; and a new southern deep water species, here named Cymbiola cooperi Petuch and Berschauer, n. sp., which ranges from Shark Bay south to near Leeman. [ABSTRACT FROM AUTHOR]

Abstract Background Anopheles vagus (subgenus Cellia) has been identified as a vector for malaria, filariasis, and Japanese encephalitis in Asia. Sporozoites of Plasmodium falciparum and Plasmodium vivax have been found in this zoophilic mosquito in Asia and Indonesia. This study systematically reviews publications regarding An. vagus species, variation, bio-ecology, and malaria transmission in various localities in Asia, especially Indonesia, to determine whether the current data support An. vagus as a species complex. Methods The databases Pubmed, Scopus, Europe PMC, and Proquest were searched to identify information regarding the morphology, karyotypes, polytene chromosome, cross-mating, ecology, and molecular identification of An. vagus was then evaluated to determine whether there were possible species complexes. Results Of the 1326 articles identified, 15 studies were considered for synthesis. The Anopheles spp. samples for this study came from Asia. Eleven studies used morphology to identify An. vagus, with singular studies using each of karyotype identification, chromosomal polytene identification, and cross-breeding experiments. Ten studies used molecular techniques to identify Anopheles spp., including An. vagus. Most studies discovered morphological variations of An. vagus either in the same or different areas and ecological settings. In this review, the members of An. vagus sensu lato grouped based on morphology (An. vagus, An. vagus vagus, An. vagus limosus, and An. limosus), karyotyping (form A and B), and molecular (An. vagus genotype A and B, An. vagus AN4 and AN5). Genetic analysis revealed a high conservation of the ITS2 fragment among members except for the An. vagus genotype B, which was, in fact, Anopheles sundaicus. This review also identified that An. vagus limosus and An. vagus vagus were nearly identical to the ITS2 sequence. Conclusion Literature review studies revealed that An. vagus is conspecific despite the distinct morphological characteristic of An. vagus and An. limosus. Further information using another barcoding tool, such as mitochondrial COI and ND6 and experimental cross-mating between the An. vagus and An. limosus may provide additional evidence for the status of An. vagus as a species complex.

Abstract Background The origin of variation is of central interest in evolutionary biology. Maternal mRNAs govern early embryogenesis in many animal species, and we investigated the possibility that heterogeneity in maternal mRNA provisioning of eggs can be modulated by environmental stimuli. Results We employed two sibling species of the ascidian Ciona, called here types A and B, that are adapted to different temperature regimes and can be hybridized. Previous study showed that hybrids using type B eggs had higher susceptibility to thermal stress than hybrids using type A eggs. We conducted transcriptome analyses of multiple single eggs from crosses using eggs of the different species to compare the effects of maternal thermal stress on heterogeneity in egg provisioning, and followed the effects across generations. We found overall decreases of heterogeneity of egg maternal mRNAs associated with maternal thermal stress. When the eggs produced by the F1 AB generation were crossed with type B sperm and the progeny (‘ABB’ generation) reared unstressed until maturation, the overall heterogeneity of the eggs produced was greater in a clutch from an individual with a heat-stressed mother compared to one from a non-heat-stressed mother. By examining individual genes, we found no consistent overall effect of thermal stress on heterogeneity of expression in genes involved in developmental buffering. In contrast, heterogeneity of expression in signaling molecules was directly affected by thermal stress. Conclusions Due to the absence of batch replicates and variation in the number of reads obtained, our conclusions are very limited. However, contrary to the predictions of bet-hedging, the results suggest that maternal thermal stress at the embryo stage is associated with reduced heterogeneity of maternal mRNA provision in the eggs subsequently produced by the stressed individual, but there is then a large increase in heterogeneity in eggs of the next generation, although itself unstressed. Despite its limitations, our study presents a proof of concept, identifying a model system, experimental approach and analytical techniques capable of providing a significant advance in understanding the impact of maternal environment on developmental heterogeneity.

Accurate species delimitation is essential for many biological subdisciplines. Nonetheless, current species diversity remains incompletely documented even in well-studied groups such as birds. Here, we take an integrative approach to examine species limits in the genus Schiffornis , a widespread group of dull-plumaged, whistle-voiced suboscine passerines of Neotropical humid-forest understory, currently considered to comprise seven species. We measured geographic variation in song, morphology, and mitochondrial and genome-wide nuclear markers to resolve the taxonomy of the genus. We show that Schiffornis comprises 13 separately evolving population lineages, of which most qualify as species taxa under all species definitions. These include a cryptic new species, several species splits, and the resurrection of a morphologically undifferentiated, but vocally and genetically distinct, taxon that was synonymized nearly a century ago in the Schiffornis turdina complex. We also found several hitherto unnoticed contact zones between diverging lineages and a leapfrog pattern of geographic song variation in the S. turdina complex, and we highlight potential avenues of further research of this genus. [ABSTRACT FROM AUTHOR]

Originally described from the northern Norway, Laonice cirrata (M. Sars, 1851) has been considered cosmopolitan and widely distributed in the North Pacific. To clarify the taxonomic status of the Pacific worms, we obtained the genetic characteristics of L. cirrata from Grøtsund Fjord, near Tromsø, one of the sites where Michael Sars collected worms to describe this species. The phylogenetic analysis of sequences of five gene fragments (mitochondrial COI and 16S rDNA, nuclear 18S rDNA and 28S rDNA, and Histone 3) showed significant difference between the Norwegian worms and worms from the north-western part of the Sea of Japan (Russia) earlier identified by morphology as L. cirrata. Common inhabitants of shallow waters in the Sea of Japan, these worms are assigned to the new species Laonice kasyanovi sp. nov. Both Northeast Atlantic and the Northwest Pacific populations exhibit high and overlapping variability of the diagnostic morphological characters of adults, and thus the two species can be considered as siblings. The distribution of these two species in the North Pacific remains uncertain and can only be elucidated by molecular data. Adults and one larva from the White Sea were also sequenced and found to be genetically identical to L. cirrata from Norway. The trochophores of L. cirrata are described and illustrated. They are characterized by two circles of large vesicles in the thick egg membrane and have been incorrectly referred to Aonides by previous authors. [ABSTRACT FROM AUTHOR]

Taxonomy, genetics and biogeography each make key contributions to biological conservation. However, integrating these disciplines to obtain a coherent account of the status of a taxon of concern not always straightforward. This is the case for the cross-border endemic plant Physaria thamnophila (Brassicaceae). This US federally-listed endangered species is restricted to a set of unique geological sites just north of the Rio Grande (Rio Bravo del Norte) river in south Texas, USA. A single highly-disjunct occurrence of this species is found on a geologically and ecologically distinct site 260 km to the south, in Tamaulipas, Mexico. In this work, we quantify the genetic differentiation between the U.S. and Mexican populations using four microsatellite markers and sequences from three nuclear genes. In both sets of data, we find a high level of genetic divergence consistent with geographic isolation on a time scale of 1-2.5 million years. Further, we provide a hypothesis for the geological basis of this geographic isolation. Integrating our data with ecological, taxonomic and conservation considerations, we propose the sub-specific designation of Physaria thamnophila subsp. loretensis for the Mexican population. The evolutionary and conservation implications of this designation are presented. [ABSTRACT FROM AUTHOR]

The article discusses the concept of cryptic species, which are morphologically similar or identical populations that are reproductively isolated. The use of molecular methods, such as DNA barcoding, has led to the identification of cryptic species in various groups of seaweeds. The article highlights a study on the biodiversity of turf-forming red algae, which revealed the presence of 12 candidate species within the Polysiphonia scopulorum complex. The authors also discuss the challenges of naming cryptic species and the importance of bringing them into a formal taxonomic framework for understanding biodiversity, invasive species, biodiscovery, and marine conservation. [Extracted from the article]

Peninsular India is an important region for mammalian diversification and harbors major biogeographic barriers. However, little is known about the role of this region in the diversification of bats though it harbors high chiropteran diversity. In this study, we used phenotypic, acoustic, and genetic markers to assess the diversification of Rhinolophus lepidus bats in South Asia. We first investigated if peninsular India is associated with speciation of R. lepidus. Further, we tested if the Palghat Gap acts as a biogeographic barrier to gene flow in this species. Our results revealed cryptic genetic diversity in peninsular India suggesting that this region holds at least one endemic species level lineage of the R. lepidus species complex.  Analyses of populations of R. lepidus across the Palghat Gap in the Western Ghats revealed clinal variation in phenotype, with bats south of this barrier being bigger and emitting echolocation calls of higher frequency. We also observed that populations on either side of the Palghat Gap have remained genetically isolated since the mid-Holocene.

Insect pests like Tribolium and Sitophilus siblings are major threats to grain storage and processing, causing quality and quantity losses that endanger food security. These closely related species, having very similar morphological and biological characteristics, often exhibit variations in biology and pesticide resistance, complicating control efforts. Accurate pest species identification is essential for effective control, but workplace safety in the grain bin associated with grain deterioration, clumping, fumigator hazards, and air quality create challenges. Therefore, there is a pressing need for an online automated detection system. In this work, we enriched the stored-grain pest sibling image dataset, which includes 25,032 annotated Tribolium samples of two species and five geographical strains from real warehouse and another 1774 from the lab. As previously demonstrated on the Sitophilus family, Convolutional Neural Networks demonstrate distinct advantages over other model architectures in detecting Tribolium. Our CNN model, MCSNet+, integrates Soft-NMS for better recall in dense object detection, a Position-Sensitive Prediction Model to handle translation issues, and anchor parameter fine-tuning for improved matching and speed. This approach significantly enhances mean Average Precision (mAP) for Sitophilus and Tribolium, reaching a minimum of 92.67 ± 1.74% and 94.27 ± 1.02%, respectively. Moreover, MCSNet+ exhibits significant improvements in prediction speed, advancing from 0.055 s/img to 0.133 s/img, and elevates the recognition rates of moving insect sibling species in real wheat storage and visible light, rising from 2.32% to 2.53%. The detection performance of the model on laboratory-captured images surpasses that of real storage facilities, with better results for Tribolium compared to Sitophilus. Although inter-strain variances are less pronounced, the model achieves acceptable detection results across different Tribolium geographical strains, with a minimum recognition rate of 82.64 ± 1.27%. In real-time monitoring videos of grain storage facilities with wheat backgrounds, the enhanced deep learning model based on Convolutional Neural Networks successfully detects and identifies closely related stored-grain pest images. This achievement provides a viable solution for establishing an online pest management system in real storage facilities. [ABSTRACT FROM AUTHOR]

The advent of genetic methods has led to the discovery of an increasing number of species that previously could not be distinguished from each other on the basis of morphological characteristics. Even though there has been an exponential growth of publications on cryptic species, such species are rarely considered in ecotoxicology. Thus, the particular question of ecological differentiation and the sensitivity of closely related cryptic species is rarely addressed. Tackling this question, however, is of key importance for evolutionary ecology, conservation biology, and, in particular, regulatory ecotoxicology. At the same time, the use of species with (known or unknown) cryptic diversity might be a reason for the lack of reproducibility of ecotoxicological experiments and implies a false extrapolation of the findings. Our critical review includes a database and literature search through which we investigated how many of the species most frequently used in ecotoxicological assessments show evidence of cryptic diversity. We found a high proportion of reports indicating overlooked species diversity, especially in invertebrates. In terrestrial and aquatic realms, at least 67% and 54% of commonly used species, respectively, were identified as cryptic species complexes. The issue is less prominent in vertebrates, in which we found evidence for cryptic species complexes in 27% of aquatic and 6.7% of terrestrial vertebrates. We further exemplified why different evolutionary histories may significantly determine cryptic species' ecology and sensitivity to pollutants. This in turn may have a major impact on the results of ecotoxicological tests and, consequently, the outcome of environmental risk assessments. Finally, we provide a brief guideline on how to deal practically with cryptic diversity in ecotoxicological studies in general and its implementation in risk assessment procedures in particular. Environ Toxicol Chem 2023;42:1889–1914. © 2023 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC. [ABSTRACT FROM AUTHOR]

A cryptic lineage is a genetically diverged but morphologically unrecognized variant of a known species. Clarifying cryptic lineage evolution is essential for quantifying species diversity. In sympatric cryptic lineage divergence compared with allopatric divergence, the forces of divergent selection and mating patterns override geographical isolation. Introgression, by supplying preadapted or neutral standing genetic variations, can promote sympatric cryptic lineage divergence via selection. However, most studies concentrated on extant species introgression, ignoring the genetic legacy of introgression from extinct or unsampled lineages ("ghost introgression"). Cycads are an ideal plant for studying the influence of ghost introgression because of their common interspecific gene flow and past high extinction rate. Here, we utilized reference‐based ddRADseq to clarify the role of ghost introgression in the evolution of a previously identified sympatric cryptic lineage in Cycas revoluta. After re‐evaluating the evolutionary independency of cryptic lineages, the group‐wise diverged single‐nucleotide polymorphisms among sympatric and allopatric lineages were compared and functionally annotated. Next, we employed an approximate Bayesian computation method for hypothesis testing to clarify the cryptic lineage evolution and ghost introgression effect. SNPs with the genomic signatures of ghost introgression were further annotated. Our results reconfirmed the evolutionary independency of cryptic lineage among C. revoluta and demonstrated that ghost introgression to the noncryptic lineage facilitated their divergence. Gene function related to heat stress and disease resistance implied ecological adaptation of the main extant populations of C. revoluta. [ABSTRACT FROM AUTHOR]

The Ceratitis FARQ species complex consists of four highly destructive agricultural pests of Africa, namely C. fasciventris , C. anonae , C. rosa , and C. quilicii. The members of the complex are considered very closely related and the species limits among them are rather obscure. Their economic significance and the need for developing biological methods for their control makes species identification within the complex an important issue, which has become clear that can only be addressed by multidisciplinary approaches. Chromosomes, both mitotic and polytene, can provide a useful tool for species characterization and phylogenetic inference among closely related dipteran species. In the current study, we present the mitotic karyotype and the polytene chromosomes of C. rosa and C. quilicii together with in situ hybridization data. We performed a comparative cytogenetic analysis among the above two species and C. fasciventris , the only other cytogenetically studied member of the FARQ complex, by comparing the mitotic complement and the banding pattern of the polytene chromosomes of each species to the others, as well as by studying the polytene chromosomes of hybrids between them. Our analysis revealed no detectable chromosomal rearrangements discriminating the three FARQ members studied, confirming their close phylogenetic relationships. [ABSTRACT FROM AUTHOR]

Background & objectives: Malaria transmission in Punjab, India is mainly seasonal with variation in its endemicity that may be due to varying vector behaviour in different areas of the state, primarily attributed to the existence of sibling species complexes among the vector species. So far there is no report regarding the existence of malaria vectors sibling species in the state of Punjab, therefore, the present study was planned to investigate the status of sibling species of two main vectors of malaria viz. Anopheles culcifacies and Anopheles fluviatilis in different districts of Punjab. Methods: Mosquito collections were made through hand catch in the morning hours. Malaria vector species An. culicifacies and An. fluviatilis were morphologically identified and man hour density was calculated. Both the vector species were subjected to molecular assays for sibling species identification through amplification of D3 domain of 28S ribosomal DNA by allele-specific PCR. Results: Four sibling species of An. culicifacies, were identified viz. A, B, C and E. Species A was identified from Bhatinda district, species B, C and E from. S.A.S. Nagar and species C from Hoshiarpur. Two sibling species S and T of An. fluviatilis were identified from districts S.A.S. Nagar and Rupnagar. Interpretation & conclusion: Presence of four sibling species of An. culicifacies and two sibling species of An. fluviatilis in Punjab necessitates planning of longitudinal studies to ascertain their role in disease transmission so that appropriate interventions may be applied to achieve malaria elimination.

The present study was carried out to design a simple, new and accurate method for identification of the morphologically similar four species (forms A and B of P. shikueii, P. mictodonta, and P. wilsoni) of the Perinereis nuntia species complex by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) patterns of the internal transcribed spacer (ITS). Restriction digestion analysis in the following two steps generated a specific restriction pattern for each species: (1) by the first digestion of amplified ITS fragments with AluI, we can distinguish the two forms of P. shikueii from P. mictodonta and P. wilsoni; (2) by the subsequent digestion of ITS fragments with Cfr10I, we can distinguish between the forms A and B of P. shikueii, and by that with HinfI, we can distinguish P. mictodonta from P. wilsoni. Our result indicates that this method enable easy and accurate identification of these taxa. We additionally examined the distributions of the two forms of P. shikueii in Japan, Korea and Taiwan using this new method. The form B was distributed in a wide range in western Japan and Korea, whereas the form A was distributed in few limited sites in the inner parts of the Seto Inland Sea and the Ariake Sea in Japan, the western and southern coasts of Korea, and the western coast of Taiwan. [ABSTRACT FROM AUTHOR]

Insect carboxylesterases (CXEs) can be expressed in multiple tissues and play crucial roles in detoxifying xenobiotic insecticides and degrading olfactory cues. Therefore, they have been considered as an important target for development of eco-friendly insect pest management strategies. Despite extensive investigation in most insect species, limited information on CXEs in sibling moth species is currently available. The Ectropis obliqua Prout and Ectropis grisescens Warren are two closely related tea geometrid species, which share the same host of tea plant but differ in geographical distribution, sex pheromone composition, and symbiotic bacteria abundance, providing an excellent mode species for studies of functional diversity of orthologous CXEs. In this study, we focused on EoblCXE14 due to its previously reported non-chemosensory organs-biased expression. First, the EoblCXE14 orthologous gene EgriCXE14 was cloned and sequence characteristics analysis showed that they share a conserved motif and phylogenetic relationship. Quantitative real-time polymerase chain reaction (qRT-PCR) was then used to compare the expression profiles between two Ectropis spp. The results showed that EoblCXE14 was predominately expressed in E. obliqua larvae, whereas EgriCXE14 was abundant in E. grisescens at multiple developmental stages. Interestingly, both orthologous CXEs were highly expressed in larval midgut, but the expression level of EoblCXE14 in E. obliqua midgut was significantly higher than that of EgriCXE14 in E. grisescens midgut. In addition, the potential effect of symbiotic bacteria Wolbachia on the CXE14 was examined. This study is the first to provide comparative expression profiles of orthologous CXE genes in two sibling geometrid moth species and the results will help further elucidate CXEs functions and identify a potential target for tea geometrid pest control. [ABSTRACT FROM AUTHOR]

Trichopoda pennipes is a tachinid parasitoid of several significant heteropteran agricultural pests, including the southern green stink bug, Nezara viridula, and leaf-footed bug, Leptoglossus phyllopus. To be used successfully as a biological control agent, the fly must selectively parasitize the target host species. Differences in the host preference of T. pennipes were assessed by assembling the nuclear and mitochondrial genomes of 38 flies reared from field-collected N. viridula and L. phyllopus. High-quality de novo draft genomes of T. pennipes were assembled using long-read sequencing. The assembly totaled 672 MB distributed among 561 contigs, having an N50 of 11.9 MB and a GC of 31.7%, with the longest contig at 28 MB. The genome was assessed for completeness using BUSCO in the Insecta dataset, resulting in a score of 99.4%, and 97.4% of the genes were single copy-loci. The mitochondrial genomes of the 38 T. pennipes flies were sequenced and compared to identify possible host-determined sibling species. The assembled circular genomes ranged from 15,345 bp to 16,390 bp and encode 22 tRNAs, two rRNAs, and 13 protein-coding genes (PCGs). There were no differences in the architecture of these genomes. Phylogenetic analyses using sequence information from 13 PCGs and the two rRNAs individually or as a combined dataset resolved the parasitoids into two distinct lineages: T. pennipes that parasitized both N. viridula and L. phyllopus, and others that parasitized only L. phyllopus. [ABSTRACT FROM AUTHOR]

The carrot cyst nematode, Heterodera carotae , is an important pest causing qualitative and quantitative production losses across several countries worldwide. In culture crops, Heterodera carotae and Heterodera cruciferae , can be found in mixture and are virtually unrecognizable from a morphological point of view. However, only H. carotae is able to develop on carrot. To monitor carrot production and set up suitable H. carotae control plans, simple and reliable diagnostic methods to detect and identify these two species are needed. In this study, we developed and successfully tested two real-time PCR protocols (SYBR Green singleplex and TaqMan multiplex real-time PCRs) against both species: H. carotae and H. cruciferae. Using two specific sets of primers, we managed, whatever the methods, to distinguish the two species. Moreover, we showed a higher specificity using the TaqMan assays but a better sensitivity with the SYBR Green assays. In this study, we are proposing two useful and reliable diagnostic tools to enable farmers and scientists to rapidly detect two morphologically and genetically closely related species and supervise agricultural management. • The cyst nematode H. carotae and H. cruciferae occur in mixture in carrot fields. • H. carotae and H. cruciferae are indistinguishable using the available molecular tools. • Two real-time PCRs were developped to distinguish H. carotae from H. cruciferae. • The two real-time PCRs were complementary in terms of sensitivity and specificity. • Microsatellite loci are usefull markers to identify species-specific polymorphisms. [ABSTRACT FROM AUTHOR]

Notiosorex is the only genus of shrews in North America with particular adaptations for arid habitats such as deserts. Five species currently are recognized in the genus, one of which, Notiosorex evotis , lives in deciduous rain forest from northern Sinaloa to the state of Jalisco in México. Notiosorex evotis originally was described as a subspecies of N. crawfordi ; however, it was recently proposed as a valid species based on a discriminant function analysis of craniodental characters. Morphological differentiation between N. evotis populations and sympatric occurrences with N. crawfordi in northern Sinaloa have been recognized. Here, we used a phylogenetic analysis of a mitochondrial gene (Cytb ; 1,140 bp) and the nuclear beta fibrinogen intron 7 (β-fib I7 ; 385 bp), as well as cranial geometric morphometrics, to assess the taxonomic status of N. evotis. We found sequences of N. evotis forming two main subclades: one that includes the populations of the state of Sinaloa, the other including populations of Nayarit and Jalisco. The boundaries between the two groups seem to be related to the ecotonal cline between the Pacific Lowlands province and the Trans-Mexican Volcanic Belt province, which acts as a geographical barrier. The discriminant function analysis revealed clear differences in skull shape between the three species of Notiosorex to the northwest of its distribution. Considering these multiple lines of evidence from our data set, we confirm that N. evotis is a monotypic species. Our results suggest that geometric morphometrics can be used successfully to identify sibling species by shape, especially in groups where determination by craniodental measurements is not possible. [ABSTRACT FROM AUTHOR]

Background: Gujarat State has been witnessing large scale urbanization, in last two decades, resulting changes in local environment and microclimate may have also influenced the resting, feeding habits and development of Anopheles cu­licifacies sensu 1ato. Therefore, a systematic longitudinal study was undertaken to know the bionomics of An. culicifa­cies s.l. in present study. Methods: The study was conducted in four sentinel villages in Kheda and Panchmahal Districts. The mosquitoes rest­ing indoors and outdoors were collected in early morning hours, using mouth aspirator, pyrethrum space spray and light traps. Mosquito landing collections on human volunteers was carried out from dusk to dawn. Species composition, abundance, seasonal prevalence, resting behavior (Endophily and Exophily), sibling species composition, vector poten­tial and insecticide susceptibility status of malaria vectors was studied. Results: Six Anopheles species were collected, An. subpictus s.l. was the predominant species followed by An. culicifa­cies s.l., a known malaria vector was resting indoor and zoophagic behaviour. Anopheles culicifacies, sibling species B (89%) was found. The sporozoite rate (%) and entomological inoculation rate in Kheda was 2.33%, 3.09 per bite/ per­son/annum and they were 1.05% and 0.475 bite/person/annum in Panchmahal, respectively. Anopheles culicifa­cies s.l. was found possible resistance to alpha-cypermethrin. Conclusion: Anopheles culicifacies s.l. showed endophillic, zoophagic behaviour and found possible resistance to al­pha-cypermethrin. Early biting behaviour of An. culicifacies s.l. in this area is a cause of concern. Therefore, there is need for frequent monitoring and evaluation of vector control measures in order to achieve the elimination target of ma­laria in this area.

Insect pests like Tribolium and Sitophilus siblings are major threats to grain storage and processing, causing quality and quantity losses that endanger food security. These closely related species, having very similar morphological and biological characteristics, often exhibit variations in biology and pesticide resistance, complicating control efforts. Accurate pest species identification is essential for effective control, but workplace safety in the grain bin associated with grain deterioration, clumping, fumigator hazards, and air quality create challenges. Therefore, there is a pressing need for an online automated detection system. In this work, we enriched the stored-grain pest sibling image dataset, which includes 25,032 annotated Tribolium samples of two species and five geographical strains from real warehouse and another 1774 from the lab. As previously demonstrated on the Sitophilus family, Convolutional Neural Networks demonstrate distinct advantages over other model architectures in detecting Tribolium. Our CNN model, MCSNet+, integrates Soft-NMS for better recall in dense object detection, a Position-Sensitive Prediction Model to handle translation issues, and anchor parameter fine-tuning for improved matching and speed. This approach significantly enhances mean Average Precision (mAP) for Sitophilus and Tribolium, reaching a minimum of 92.67 ± 1.74% and 94.27 ± 1.02%, respectively. Moreover, MCSNet+ exhibits significant improvements in prediction speed, advancing from 0.055 s/img to 0.133 s/img, and elevates the recognition rates of moving insect sibling species in real wheat storage and visible light, rising from 2.32% to 2.53%. The detection performance of the model on laboratory-captured images surpasses that of real storage facilities, with better results for Tribolium compared to Sitophilus. Although inter-strain variances are less pronounced, the model achieves acceptable detection results across different Tribolium geographical strains, with a minimum recognition rate of 82.64 ± 1.27%. In real-time monitoring videos of grain storage facilities with wheat backgrounds, the enhanced deep learning model based on Convolutional Neural Networks successfully detects and identifies closely related stored-grain pest images. This achievement provides a viable solution for establishing an online pest management system in real storage facilities.

Species Chironomus sp. prope agilis Kiknadze, Siirin, Filippova et al., 1991 belongs to the Ch. plumosus group of sibling species. It was described on the basis of its karyotype and analysis of isozymes from one population in the Urals but since then no quantitative data on chromosomal polymorphism of this species have been published. The goal of this study is to broaden our knowledge of the chromosomal polymorphism and distribution of the Chironomus sp. prope agilis, which, along with the data on chromosomal polymorphism of other species from the Ch. plumosus group, can give us a better understanding of the connection between chromosomal polymorphism and ecological conditions of habitats. The specimens of Chironomus sp. prope agilis were found only in 8 natural populations from the Urals, Western Siberia and Kazakhstan, which allows us to conclude that the species range of Chironomus sp. prope agilis is not as wide as for most other species from Ch. plumosus group. An analysis of chromosomal polymorphism in these 8 natural populations of Chironomus sp. prope agilis has been performed. All of the studied populations were either monomorphic or showed very low level of chromosomal polymorphism, with 4.4-8.7% of heterozygous specimens per population and 0.04-0.08 heterozygotic inversion per larvae. The total number of banding sequences found in the banding sequence pool of Chironomus sp. prope agilis is 10. The mapping of banding sequence p'ag2B3 is presented for the first time. Besides inversions, one reciprocal translocation was found in a population from Kazakhstan, B-chromosome was found in one population from the Urals region of Russia, and heterozygosity of the level of expression of Balbiany rings in arm G was observed in several studied populations. [ABSTRACT FROM AUTHOR]

We used three-dimensional micro-computed tomography to compare the skeletal features of two species of male tree frogs with similar morphologies: the globally prevalent Hyla japonica and critically endangered Hyla suweonensis. One of the most prominent differences between the two species was the angle of the alary processes of the premaxilla. In H. suweonensis, the alary processes were inclined posteriorly at an angle of 77.05°, whereas in H. japonica, they were deflected at an angle of 93.27° (a difference of more than 16°). This difference accounts for the more protruded shape of H. suweonensis compared with that of H. japonica. Sizes and proportions related to skeletal compartments (e.g. skull, forelimb, vertebrae, and hindlimb) also differed between the two species, which is consistent with the general exterior observation that compared with H. japonica, H. suweonensis has a more acuminate snout and a more slender body. The full skeletal configuration could provide classification keys for the two sympatric species that will be applicable to ecological field surveys and conservation activities.

Although molecular methods and bioacoustical analysis have been used to uncover cryptic species, the combination of both methodologies is still rare. The humbug damselfish complex, Dascyllus aruanus , is composed of at least two species with Dascyllus aruanus in the Pacific Ocean and Dascyllus abudafur in the Indian Ocean. However, genetic data suggest that additional species could be found. Therefore, we aimed to determine whether the sounds produced by different populations of fish could help to distinguish cryptic species. Recordings of chase and courtship sounds were made on humbug damselfish populations from Madagascar, Taiwan and French Polynesia. Chase sound features are more variable than courtship sounds, suggesting more constraints on courtship sounds, since they would contribute to premating isolation. Comparison between courtship sounds show the variation in acoustic features between Taiwan and Madagascar align with genetic differences, supporting that sounds could discriminate cryptic species in Teleosts. Moreover, differences in both acoustic features and genetic data are also found between Taiwan and French Polynesia, suggesting two clearly distinct populations. Consequently, the name D. emamo can be resurrected for the Polynesian humbug damselfish. External phenotype traits do not allow the distinction between populations, illustrating that only behaviour has been modified. [ABSTRACT FROM AUTHOR]

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The flour beetle Tribolium freemani is a sibling species of the model organism and important pest Tribolium castaneum. The two species are so closely related that they can produce hybrid progeny, but the genetic basis of their differences has not been revealed. In this work, we sequenced the T. freemani genome by applying PacBio HiFi technology. Using the well-assembled T. castaneum genome as a reference, we assembled 262 Mb of the T. freemani genomic sequence and anchored it in 10 linkage groups corresponding to nine autosomes and sex chromosome X. The assembly showed 99.8% completeness of conserved insect genes, indicating a high-quality reference genome. Comparison with the T. castaneum assembly revealed that the main differences in genomic sequence between the two sibling species come from repetitive DNA, including interspersed and tandem repeats. In this work, we also provided the complete assembled mitochondrial genome of T. freemani. Although the genome assembly needs to be ameliorated in tandemly repeated regions, the first version of the T. freemani reference genome and the complete mitogenome presented here represent useful resources for comparative evolutionary studies of related species and for further basic and applied research on different biological aspects of economically important pests. [ABSTRACT FROM AUTHOR]

Background: Gujarat State has been witnessing large scale urbanization, in last two decades, resulting changes in local environment and microclimate may have also influenced the resting, feeding habits and development of Anopheles culicifacies sensu 1ato. Therefore, a systematic longitudinal study was undertaken to know the bionomics of An. culicifacies s.l. in present study. Methods: The study was conducted in four sentinel villages in Kheda and Panchmahal Districts. The mosquitoes resting indoors and outdoors were collected in early morning hours, using mouth aspirator, pyrethrum space spray and light traps. Mosquito landing collections on human volunteers was carried out from dusk to dawn. Species composition, abundance, seasonal prevalence, resting behavior (Endophily and Exophily), sibling species composition, vector potential and insecticide susceptibility status of malaria vectors was studied. Results: Six Anopheles species were collected, An. subpictus s.l. was the predominant species followed by An. culicifacies s.l., a known malaria vector was resting indoor and zoophagic behaviour. Anopheles culicifacies, sibling species B (89%) was found. The sporozoite rate (%) and entomological inoculation rate in Kheda was 2.33%, 3.09 per bite/ person/ annum and they were 1.05% and 0.475 bite/person/annum in Panchmahal, respectively. Anopheles culicifacies s.l. was found possible resistance to alpha-cypermethrin. Conclusion: Anopheles culicifacies s.l. showed endophillic, zoophagic behaviour and found possible resistance to alpha- cypermethrin. Early biting behaviour of An. culicifacies s.l. in this area is a cause of concern. Therefore, there is need for frequent monitoring and evaluation of vector control measures in order to achieve the elimination target of malaria in this area. [ABSTRACT FROM AUTHOR]

The endobenthic deposit-feeder polychaetes of the Capitella capitata species complex have a strong relation with organically enriched sediments (OES). The effect of non-organically enriched sediments (NOES, 0.09% of organic content) and OES (6.26% of organic content) on mortality, growth and development of juveniles and adults of Capitella teleta (cultured for more than 40 years) and Capitella sp. from Alfacs Bay, Tarragona, Spain (acclimatized for one month) were analysed in laboratory conditions. Juveniles and adults were exposed (all individuals separately in different dishes) to the two treatments, periodically observed and measured until death. NOES considerably impaired survival, growth rates and development of juveniles and adults of both species, which showed highly variable negative growth rates under OES. Juvenile and adult C. teleta showed an improved maturity under OES, even producing two protandrous and one protogynous individuals, whereas no Capitella sp. reached maturity during the experiment.

Composting earthworms of the genus Eisenia play an important role in soil ecosystems. However, taxonomic classification of this genus, especially the sibling species Eisenia fetida and Eisenia andrei, is complicated because of their morphological similarity. In this study, we assessed the utility of the complete mitochondrial genome (mitogenome) for identification and differentiation of the two species. The complete mitogenomes of E. andrei and E. fetida were 15,714 and 16,560 bp, respectively. They contained 37 genes, comprising 13 protein-coding genes (PCGs), two rRNA genes, 22 tRNA genes, and a putative noncoding region, as observed in other earthworms. Sequence comparisons based on the complete nucleotide sequences excluding the non-coding region showed 85.8% similarity, whereas the predicted amino acid sequences of the 13 PCGs were 92.7% similar between the two species. In particular, distinct features were found in the non-coding regions of the mitogenomes. They include a control region associated with putative mitogenome replication and an extended sequence. The extended sequence showed significant differences between the two species and other known earthworm species, suggesting its potential as a feasible molecular marker for species identification. Phylogenetic analysis of the 36 mitogenomes of earthworm species corroborated the monophyly of the genus Eisenia and the taxonomic distinctness of the sibling species pair, E. fetida and E. andrei. [ABSTRACT FROM AUTHOR]

The West-Palaearctic Colobopsis ant populations have long been considered a single species (Colobopsis truncata). We studied the diversity of this species by employing a multidisciplinary approach and combining data from our surveys, museum and private collections, and citizen science platforms. As a result, we have revealed the existence of a second species, which we describe as Colobopsis imitans sp. nov. , distributed allopatrically from Co. truncata and living in the Maghreb, Sicily and southern Iberia. While the pigmentation of Co. truncata is reminiscent of Dolichoderus quadripunctatus , that of Co. imitans is similar to Crematogaster scutellaris , with which Co. imitans lives in close spatial association, and whose foraging trails it habitually follows, similar to Camponotus lateralis and other ant-mimicking ants. The isolation between Co. imitans and Co. truncata seems to have occurred relatively recently because of significant, yet not extreme, morphometric differentiation, and to mtDNA polyphyly. Both Co. imitans and Co. truncata appear to employ mimicry of an unpalatable or aggressive ant species as an important defensive strategy; this 'choice' of a different model species is motivated by biogeographic reasons and appears to act as a critical evolutionary driver of their diversification. [ABSTRACT FROM AUTHOR]

In this study, Anopheles mosquitoes were characterized through an investigation of adult female mosquitoes for sibling species. For the first time, researchers combined efforts on morphology and DNA barcoding utilizing the markers cytochrome c oxidase subunit I (COI) gene and internal transcribed spacer 2 (ITS2) region was made. Cytochrome c oxidase subunit I and ITS2 regions of morphologically differentiated, Anopheline mosquitoes were sequenced. According to the results of molecular identification (PCR) of the distinctive sibling species of the Anopheline mosquitoes, two sibling species from An. gambiae complex and An. rufipes group were identified viz; An. gambiae ss (Giles 1902) and An. rufipes (Gough 1910), these sibling species were discovered in Bosso LGA's Large water and Gutter. An. gambiae s.s was collected in all the breeding habitats sampled in Katcha, Lapai and Shiroro LGAs. In this report, the habitat breeding density of the An. gambiae s.s. was found to be higher compared to An. rufipes, which was restricted to large water and gutter breeding habitats. This study highlights the usefulness and feasibility of COI and ITS2 genetic markers in recognizing Anophelines, their sibling species, the importance of a unified, systematic approach in mosquito taxonomy and highlights the epidemiological role of An. gambiae s.s. For the first time in Niger State An. rufipes has appeared as a malaria vector, emphasising the need for proper monitoring of this species across the country to effectively set up vector management strategies. [ABSTRACT FROM AUTHOR]

Distribution of sibling species Larus cachinnans (Pallas, 1811) and Larus michahellis (J. F. Naumann 1840) were studied on coast of the Black Sea. Originally, Larus michahellis breeds on seaside cliffs and rocky islets of the Southern part of the Black Sea and of the Crimean Peninsula. Having adapted to breeding on the roofs of buildings, this species is spread to the north along the seaside: to Novorossiysk on the eastern coast, to Constanta on the western one. Additionally, an isolated population of Larus cachinnans breeds on low-lying islands and sand spits mostly in the Northwestern Black Sea. Here, the species spreads on the seaside from Constanta northward to Karkinitian Bay. Further to the east, breeding populations of Caspian Gull are found on the Black Sea coast only at the southeastern edge of the Taman Peninsula to the south to Anapa. The coast of the Sea of Azov, adjacent to the northeastern part of the Black Sea, is densely populated by Larus cachinnans. Only in the southern part of the Sea of Azov, on the coast of the Kerch Peninsula, there is a small population of Larus michahellis. [ABSTRACT FROM AUTHOR]

Abstract Background Anopheles subpictus and Anopheles sundaicus are closely related species, each comprising several sibling species. Ambiguities exist in the classification of these two nominal species and the specific status of members of these species complexes. Identifying fixed molecular forms and mapping their spatial distribution will help in resolving the taxonomic ambiguities and understanding their relative epidemiological significance. Methods DNA sequencing of Internal Transcribed Spacer-2 (ITS2), 28S-rDNA (D1-to-D3 domains) and cytochrome oxidase-II (COII) of morphologically identified specimens of two nominal species, An. subpictus sensu lato (s.l.) and An. sundaicus s.l., collected from the Indian subcontinent, was performed and subjected to genetic distance and molecular phylogenetic analyses. Results Molecular characterization of mosquitoes for rDNA revealed the presence of two molecular forms of An. sundaicus s.l. and three molecular forms of An. subpictus s.l. (provisionally designated as Form A, B and C) in the Indian subcontinent. Phylogenetic analyses revealed two distinct clades: (i) subpictus clade, with a single molecular form of An. subpictus (Form A) prevalent in mainland India and Sri Lanka, and (ii) sundaicus clade, comprising of members of Sundaicus Complex, two molecular forms of An. subpictus s.l. (Form B and C), prevalent in coastal areas or islands in Indian subcontinent, and molecular forms of An. subpictus s.l. reported from Thailand and Indonesia. Based on the number of float-ridges on eggs, all An. subpictus molecular Form B were classified as Species B whereas majority (80%) of the molecular Form A were classified as sibling species C. Fixed intragenomic sequence variation in ITS2 with the presence of two haplotypes was found in molecular Form A throughout its distribution. Conclusion A total of three molecular forms of An. subpictus s.l. and two molecular forms of An. sundaicus s.l. were recorded in the Indian subcontinent. Phylogenetically, two forms of An. subpictus s.l. (Form B and C) prevalent in coastal areas or islands in the Indian subcontinent and molecular forms reported from Southeast Asia are members of Sundaicus Complex. Molecular Form A of An. subpictus is distantly related to all other forms and deserve a distinct specific status.

The correct identification of single specimens on a particular area has great importance in establishing appropriate biodiversity protection programs. Species of the genus Merodon Meigen, 1803 (Diptera, Syrphidae) represent important pollinators that are particularly associated with the pollination of wild and cultivated bulbous plants, both wild and cultivated. In order to contribute to a taxonomic issue of separating two cryptic, sibling hoverfly species of M. avidus species complex, we programmed and trained specific prediction model that was able to specify to which of two assumed species (M. avidus or M. moenium) each database specimen belongs. Using two ML techniques (artificial neural network (ANN) and gradient boosting machine (GBM)), we created two separable models, depending on a variable used for a prediction (Model 1 - modelling based on a geographic variable, Model 2 - modelling based on a temporal variable). Moreover, each model was trained and tested with different data sets, resulting in a different predictive accuracy. While ANN modelling showed a higher percent of correct determination when using surrogate information than when using reduced (basic) data set, GBM modelling has given a quite stable result through all three data types. In both ML approaches, comparing Model 1 and Model 2 results showed that prediction based on a temporal variable (day, month and a year of specimen sampling) reached a better predictive performance than a prediction based on a longitude and latitude, on all data sets. This led us to the conclusion that information about the time of sampling was more useful for creating desired determination key with artificial intelligence algorithms than information about longitude and latitude of sampling localities. Therefore, we suggest that time of activity of adult specimens could have been of greater importance in the differentiation of M. avidus and M. moenium species from a common ancestor. The environmental factors and selective forces connected with the season might have had a more important role in M. avidus / M. moenium speciation, compared to environmental factors / selective pressures connected with the geographic position of their activity. The demonstrated modelling represents a positive signal in the field of potential implementation of these systems as support in the initial determination Merodon specimens. We suggest it2s potential use as technical support in old and partially unreliable databases, in determination of fresh sampled specimens as well as in finding the most efficient sampling strategies.

Abstract Background Anopheles fluviatilis is a species-complex comprising of four cryptic species provisionally designated as species S, T, U and V. Earlier, a 28S-rDNA based allele-specific polymerase chain reaction (ASPCR) assay was developed for the differentiation of the then known three members of the An. fluviatilis complex, i.e., species S, T, and U. This assay was modified in consequence of the discovery of a new cryptic member, species V, in the Fluviatilis Complex to include identification of new species. Methods In the modified procedure, the ASPCR assay was performed first, followed by restriction digestion of PCR product with an enzyme BamH I, which cleaves specifically PCR amplicon of species V and the resultant PCR–RFLP products can differentiate all the four cryptic members of the complex. Morphologically identified An. fluviatilis samples were subjected to sibling species identification by modified PCR-based assay and standard cytotaxonomy. The result of PCR-based assay was validated through cytotaxonomy as well as DNA sequencing of some representative samples. Results The modified PCR-based assay differentiates all four sibling species. The result of modified PCR-based assay tested on field samples was in agreement with results of cytotaxonomy as well as DNA sequencing of representative samples. Conclusions The modified PCR-based assay unambiguously differentiates all four known members of the An. fluviatilis species complex. This assay will be useful in studies related to bionomics of members of the Fluviatilis Complex in their role in malaria transmission.

Distribution of sibling species Larus cachinnans (Pallas, 1811) and Larus michahellis (J. F. Naumann 1840) were studied on coast of the Black Sea. Originally, Larus michahellis breeds on seaside cliffs and rocky islets of the Southern part of the Black Sea and of the Crimean Peninsula. Having adapted to breeding on the roofs of buildings, this species is spread to the north along the seaside: to Novorossiysk on the eastern coast, to Constanta on the western one. Additionally, an isolated population of Larus cachinnans breeds on low-lying islands and sand spits mostly in the Northwestern Black Sea. Here, the species spreads on the seaside from Constanta northward to Karkinitian Bay. Further to the east, breeding populations of Caspian Gull are found on the Black Sea coast only at the southeastern edge of the Taman Peninsula to the south to Anapa. The coast of the Sea of Azov, adjacent to the northeastern part of the Black Sea, is densely populated by Larus cachinnans. Only in the southern part of the Sea of Azov, on the coast of the Kerch Peninsula, there is a small population of Larus michahellis.

We document a ~700 km northward range extension of the rare Andaman grunt, Pomadasys andamanensis McKay and Satapoomin 1994, to the Bangladesh coast of the north-eastern Bay of Bengal. Sixteen specimens (82-129 mm SL) were collected from fishermen catches in Ukhia, Teknaf and Zinjira Island, south-eastern Bangladesh, during 2014-2019 and their counts, measurements and descriptions are provided in detail. In addition, underwater videography (near Zinjira Island), shows an individual swimming over sand-gravel-rock bottom covered with a dense bed of brown algae, predominantly Padina and Dictyota. Using the Cytochrome Oxidase subunit I (COI) region sequence variation of mitochondrial DNA, four specimens were found genetically indistinguishable from the topotype (Andaman Sea coast of Thailand) specimens collected in this study. Long regarded as an endemic species to the Andaman Sea (type locality), the distribution range of P. andamanensis now extends from Phuket Island in the south to the Inani Coast in Cox's Bazar, Bangladesh in the north, spanning roughly 1500 km in the Andaman Sea and eastern Bay of Bengal. Comparison between published COI barcode data of P. andamanensis and the barcodes generated in this study indicates the presence of a cryptic sibling species from southern Bali, Indonesia, in the southwestern end of the Coral Triangle. A deeper phylogenetic and taxonomic investigation covering more Pomadasys spp. in the Bay of Bengal and neighboring region is suggested to resolve species level ambiguities. [ABSTRACT FROM AUTHOR]

It is shown that under the old concept of Platyja cyanocraspis Hampson, 1922 at least six species are intermingled, including two sympatric sibling ones from New Guinea. Four such species are new to science and are herein described, namely Platyja subtracta sp. n. (New Guinea), P. togutila sp. n. (Halmahera), P. vityaz sp. n. (Bismarck Archipelago) and P. yaleyambae sp. n. (Louisiades), while a fifth one from Buru and Ceram is upgraded from subspecific status as P. lecerfi A.E. Prout, 1922 stat. n. A singleton from New Britain (Bismarck Archipelago) closely albeit not entirely matching P. vityaz is also discussed.

The black-throated trogon, Trogon rufus , is a widespread, polytypic species-complex with a convoluted taxonomic history. Here, we integrated morphological, vocal and genetic datasets, including spectral data and digital quantification of barred plumage, to assess and redefine its species limits according to the foremost species concepts. We suggest the recognition of four named and one new species. Trogon tenellus and T. cupreicauda are divergent across Central and South America without geographic overlap or intermediates. Trogon chrysochloros in the Atlantic Forests of Brazil is phenotypically, genetically and ecologically distinct. In Amazonia, Trogon rufus consists of three phenotypically distinct subspecies intergrading with each other in a ring-like formation around central Amazonian rivers. Trogon rufus rufus in the Guiana Shield, Trogon rufus amazonicus in south-eastern Amazonia and Trogon rufus sulphureus in western Amazonia, with contact across the Lower Amazon and Madeira rivers, likely due to secondary contact between incompletely diverged lineages. The unique combination of song, morphology and mtDNA features of an unnamed, isolated population in the Atlantic Forest of north-eastern Brazil resulted in its description as a new species, known only from the type locality and considered here as Critically Endangered, requiring urgent conservation actions. [ABSTRACT FROM AUTHOR]

Abstract India has committed to eliminate malaria by 2030. The national framework for malaria elimination released by the Government of India plans to achieve this goal through strategic planning in a phased manner. Since vector control is a major component of disease management and vector elimination, it requires a thorough understanding of the biology and bionomics of malaria vectors exhibiting definite distribution patterns in diverse ecosystems in the country. Although a wealth of information is available on these aspects, lesser-known data are on biting time and rhythm, and the magnitude of outdoor transmission by the vectors which are crucial for effective implementation of the key vector control interventions. Most of the data available for the vector species are at sensu lato level, while the major vectors are species complexes and their members distinctly differ in biological characters. Furthermore, the persistent use of insecticides in indoor residual spray and long-lasting insecticidal nets has resulted in widespread resistance in vectors and changes in their behaviour. In this document, challenges in vector control in the Indian context have been identified and possible solutions to overcome the problem are suggested. Adequate addressing of the issues raised would greatly help make a deep dent in malaria transmission and consequently result in disease elimination within the targeted time frame.